Chronic or intermittent sleep disorders including narcolepsy, REM behavior disorder, sleep apnea, and insomnia afflict nearly 50-70 million people in the United States. Yet the neural mechanisms controlling both normal sleep and its pathologies remain poorly understood. Considerable evidence indicates that mesopontine cholinergic neurons and other neurons at the mesopontine junction are critical for this control and that their disregulation is involved in narcolepsy, REM behavior disorder, Parkinson's disease, supranuclear palsy and depression. The long term goal of this project is to understand the mechanisms regulating activity of these neurons and their functions in regulating sleep and sleep pathologies. Compelling evidence indicates that disruption of the Hypocretin/Orexin (Hcrt/Orx) peptide system results in narcolepsy - a sleep disorder characterized by excessive daytime sleepiness, sleep fragmentation and the intrusion of rapid eye movement sleep behaviors into wakefulness. Building on the findings from previous funding periods, which showed that mesopontine cholinergic neurons and dorsal raphe serotonergic neurons are important targets of these peptides, we will continue to investigate the general hypothesis that Hcrt/Orx peptides regulate both the short- term and long-term excitability of these neurons. We will explicitly test the hypothesis that cholinergic transmission resulting from mesopontine cholinergic neurons contribute the disruptions of waking and sleep in narcoleptic mice and that they promote the entrance into cataplexy but that their activity contributes little to regulating te duration of cataplectic attacks. To do so we will 1) identify the mechanisms underlying a new class of post-synaptic orexin actions post-spike afterhyperpolarization in mesopontine cholinergic neurons and dorsal raphe serotonergic neurons. 2) Explore the impact of the major orexin actions on firing pattern and encoding of inputs by mesopontine cholinergic neurons and dorsal raphe serotonergic neurons. 3) determine whether selective excitation and inhibition of mesopontine cholinergic neurons can regulate arousal and sleep in normal and narcoleptic mice and whether these neurons can trigger cataplexy in narcoleptic mice. These experiments will use whole-cell patch clamp recording, calcium imaging and dynamic clamp methods in brain slices from normal and orexin receptor knockout mice and will utilize optogenetic stimulation methods in normal and narcoleptic mice. Collectively, these results will advance our understanding of the molecular and cellular mechanisms underlying sleep regulation and its pathology.